Detection of Colletotrichum coccodes from soil and potato tubers by conventional and quantitative real-time PCR

Colletotrichum coccodes is the causal agent of the potato blemish disease black dot. Two PCR primer sets were designed to sequences of the ribosomal internal transcribed spacer (ITS1 and ITS2) regions for use in a nested PCR. The genus-specific outer primers (Cc1F1/Cc2R1) were designed to regions common to Colletotrichum spp., and the species-specific nested primers (Cc1NF1/Cc2NR1) were designed to sequences unique to C . coccodes . The primer sets amplified single products of 447 bp (Cc1F1/Cc2R1) and 349 bp (Cc1NF1/Cc2NR1) with DNA extracted from 33 European and North American isolates of C. coccodes. The specificity of primers Cc1NF1/Cc2NR1 was confirmed by the absence of amplified product with DNA of other species representing the six phylogenetic groups of the genus Colletotrichum and 46 other eukaryotic and prokaryotic plant pathogenic species. A rapid procedure for the direct extraction of DNA from soil and potato tubers was used to verify the PCR assay for detecting C. coccodes in environmental samples. The limit of sensitivity of PCR for the specific detection of C. coccodes when inoculum was added to soils was 3·0 spores per g, or the equivalent of 0·06 microsclerotia per g soil, the lowest level of inoculum tested. Colletotrichum coccodes was also detected by PCR in naturally infested soil and from both potato peel and peel extract from infected and apparently healthy tubers. Specific primers and a TaqMan fluorogenic probe were designed to perform quantitative real-time (TaqMan) PCR to obtain the same levels of sensitivity for detection of C. coccodes in soil and tubers during a first-round PCR as with conventional nested PCR and gel electrophoresis. This rapid and quantitative PCR diagnostic assay allows an accurate estimation of tuber and soil contamination by C. coccodes .     

Combined effect of generally regarded as safe (GRAS) compounds andTrichoderma harzianum on the control of postharvest diseases of rambutan

Botryodiplodia theobromae, Colletotrichum gloeosporioides andGliocephalotrichum microchlamydosporum are the causal fungi of the rambutan postharvest diseases stem-end rot, anthracnose and brown spot, respectively. Two different treatments of rambutan fruits(Nephelium lappaceum) against the three pathogens were compared: potassium metabisulphite (250 ppm) or cinnamaldehyde (30 ppm), each combined withTrichoderma harzianum (TrH 40). The application of TrH 40 and potassium metabisulphite effectively controlled the incidence and severity of the three postharvest diseases and maintained the overall quality and color of the fruit under low temperature storage at 13.5°C and 95% r.h. for 18 days. The greatest effect of this treatment was shown onG. microchlamydosporum. Cinnamaldehyde affected the growth and germination of TrH 40, whereas potassium metabisulphite did not. http://www.phytoparasitica.org posting Nov. 4, 2001.     

烟草低头黑病及其治方法研究进展

烟草低头黑病是烟草大田生长期为害较重的一种病害。近年来,由于烟田连作和地下害虫的发生,烟草低头黑病的发生程度和面积逐年上升。阐述了烟草低头黑病的发病症状、病原、致病性、流行规律以及综合防治方法。     

橡胶树炭疽病生防内生菌的分离鉴定及抑菌作用研究

目前化学农药的缺点日益突显,研发环保的生防菌剂是当前绿色安全的一种方法,筛选具有抑制橡胶树炭疽病的生防菌株,为研发新型生防菌剂储备资源。本文采用组织培养法分离纯化橡胶树内生菌,平板对峙筛选拮抗菌株;通过菌株培养特征、生理生化、根据16S rRNA和gyrA基因序列构建系统发育树确定其分类地位;采用平板对峙法研究内生菌株的抗菌持久性、稳定性、广谱性;采用牛津杯法研究拮抗内生菌对杀菌剂的敏感性。从橡胶树健康的根组织中分离得到4株对橡胶树炭疽病具有抑制作用的细菌（Bac RZS3D4-1、Bac RZS3D4-2、Bac RZS3D4-3、Bac RZS3D4-4）,系统发育树分析结果显示这4株细菌与解淀粉芽孢杆菌（Bacillus amyloliquefaciens）亲缘关系很近,且培养性状和生理生化特征也与解淀粉芽孢杆菌相似。这4株内生菌的抗菌持久性强,抗菌稳定性好;对尖孢炭疽菌（Colletotrichum acutatum）DFMP1E和MLZZP3、小孢拟盘多毛孢（Pestalotiopsis microspora）MF375898均有较强的抑菌能力,对茄类镰刀菌（Fusarium solani）XJ160901的抑制作用较弱;对多菌灵、异菌脲、百菌清、苯醚甲环唑4种杀菌剂均不敏感,有望开发为生防菌剂。     

Isolation and pathogenicity of Colletotrichum spp. causing anthracnose of strawberry in south west Spain

Anthracnose, caused by Colletotrichum spp., is a major disease of cultivated strawberry, Fragaria × ananassa. This study identifies the Colletotrichum spp. which causes strawberry anthracnose in the southwest of Spain. A survey of the region was carried out, and the strains isolated were identified as C. acutatum by using the polymerase chain reaction (PCR) with genus and species-specific primers, demonstrating that this species is currently the causal agent of strawberry anthracnose in the studied region. The pathogenicity of C. acutatum and C. gloeosporioides strains was evaluated on two principal strawberry cultivars (cvs Camarosa and Ventana) under field conditions, the latter being more pathogenic than the former.     

垂序商陆及胜红蓟提取物对降香黄檀炭疽病菌的抑菌机理研究

通过测定垂序商陆与胜红蓟乙酸乙酯提取物处理后的降香黄檀炭疽病菌内丙二醛含量、蛋白质含量以及4种酶活性,来初步判断垂序商陆与胜红蓟提取物对炭疽病菌的作用机理。结果表明:两种提取物都能在一定程度上破坏炭疽病病原菌膜系统,垂序商陆处理组丙二醛含量在56h后显著提高,胜红蓟处理组则缓慢增加,略高于对照组;垂序商陆处理组菌体内蛋白质含量先增加,32h后急剧降低,胜红蓟处理组蛋白质含量从初期开始就少于对照组;两种提取物都能影响菌体的保护性酶活性,两处理组SOD活力都呈现先增高后降低的趋势,CAT活力高于对照组。两组SDH、LDH活力都小于对照组,而垂序商陆处理组SDH、LDH活力小于胜红蓟处理组。初步说明两种植物提取物可通过影响病原菌的代谢系统和生理过程来发挥其抑菌作用。     

墨兰炭疽病菌生物学特性研究

以墨兰炭疽病菌为试材,研究墨兰炭疽病菌的生物学特性。结果表明:菌丝生长和孢子萌发最适温度为26℃,产生孢子最适温度为30℃;菌丝生长最适pH为7～8,产生孢子最适pH为7～9,孢子萌发最适pH为7;26℃下连续光照产孢量最多;相对湿度低于81%时分生孢子不能萌发。     

江西瑞昌山药炭疽病菌的形态及分子鉴定

为了明确江西瑞昌山药炭疽病病原菌种类归属,本文从当地采集呈典型症状的炭疽病叶片进行了病原菌分离鉴定.通过组织分离获得8个在培养性状和分生孢子形态大小均一致且均具有致病性的分离株,8个分离株在PDA平板上菌落初为白色,后变为灰色至深灰色,菌落中央产生橘红色黏质分生孢子团.分生孢子无色,长椭圆形至纺锤形,单胞,大小为(15.6~18.0)μm×(3.6~6.0)μm.对其中之一的分离株YRRC-1进行rDNA-ITS区段扩增和序列测定,获得长度为536 bp的rDNA-ITS序列,该序列与胶孢炭疽菌(Colletotrichum gloeosporioides)的对应序列同源性达100%.根据分离病菌的培养特征、形态大小和序列鉴定结果,认为瑞昌山药炭疽病菌属于胶孢炭疽菌(C. gloeosporioides).     

油茶炭疽菌携带病毒种类鉴定及多样性分析

 炭疽菌是重要的植物病原真菌,能够引起多种植物炭疽病。真菌病毒广泛存在于植物病原菌中,侵染炭疽菌属真菌的病毒资源有待挖掘。本文利用宏病毒组测序技术,对分离自我国南部6个省份不同油茶园的40株油茶炭疽菌中的病毒进行了鉴定。经过同源比对分析,发现10种基因组类型均为正义单琏RNA的新病毒。它们分属于减毒病毒科(Hypoviridae)、裸露病毒科(Narnaviridae)、线粒体病毒科(Mitoviridae)和葡萄孢欧尔密病毒科(Botourmiaviridae)。本研究是我国南方油茶炭疽病菌携带病毒的首次报道。研究结果丰富了炭疽菌属所携带的病毒基因组信息,可为深入分析炭疽菌真菌病毒的多样性和分子特性奠定基础。     

金毛狗脊内生细菌分离鉴定及其拮抗作用分析

为分离药用珍稀植物金毛狗脊内生细菌,筛选出对滇黄精上分离到的胶孢炭疽菌和腐皮镰刀菌病菌抑制效果明显的菌株。采用平板对峙法筛选拮抗菌株,通过生理生化反应及16 S rDNA序列分析,结合伯杰氏细菌鉴定手册,鉴定具有明显抑菌效果的菌株。结果表明：从金毛狗脊中分离获得31株内生细菌,菌株JMGJ01对胶孢炭疽菌和腐皮镰刀菌病菌有明显的拮抗作用,鉴定出JMGJ01菌株属于类芽孢杆菌属。